Project Summary/Abstract In contrast to typical Friedreich's ataxia (FRDA) patients with GAA expansions on both frataxin (FXN) alleles, there are some patients who carry a GAA expansion on one allele and a missense mutation on the other allele. What's most interesting about these patients is the observed phenotypic variability, which may be explained by characterizing the distinct effects these mutations have on FXN. Mitochondria processing peptidase (MPP) processes FXN to its mature functional form, which is necessary for proper mitochondria function. In typical FRDA patients, mature FXN levels and disease severity correlates with GAA expansion length. In patients with point mutations, phenotype is less readily predicted, and is mutation selective for unclear reasons. My long-term goals are to define how mutations in FRDA lead to phenotypic variability and identify therapeutic strategies to overcome this. Preliminary data revealed FRDA-associated missense mutations have distinct effects on FXN localization to the mitochondria, protein levels, and protein processing. FXNI154F, FXNG130V, and FXNW168R show impaired processing from intermediate to mature form as evidenced by increased intermediate to mature ratio and intermediate insoluble to soluble ratio. I will first examine the mechanism by which these mutations impair FXN processing by examining the interactions between FXN and MPP by co-immunoprecipitation and site-directed mutagenesis of the MPP cleavage site that produces the intermediate form. Next, I will define how G130V influences FXN's role in cellular metabolism and cellular phenotype. While typical FRDA patients carry less than 10% of control FXN levels, patients carrying the G130V missense mutation, interestingly, have less than 5 % of control mature FXN levels, yet present with a very mild phenotype. Platelets from typical FRDA patients exhibit decreased glucose incorporation and utilization compared to controls suggesting metabolic abnormalities associated with disease, and a potential way to measure genotype-phenotype correlations. Metabolic intermediates will be labeled and measured to track cellular metabolic activity and identify metabolic abnormalities between typical FRDA patients, patients with G130V and controls. These levels will then be compared following application of fatty acids as a therapeutic strategy to restore metabolism and cell phenotype. These studies will provide insight to better understanding phenotypic variability in FRDA by characterizing the distinct effects each mutation has on FXN, as well as identify new therapeutic targets for gene therapy and drug development. Additionally, this work will advance basic understanding of the function and role of FXN, as it necessary for proper mitochondria function to support both the normal and diseased nervous system, and provide understanding for other neurological disorders, which will lead to the development of better treatment and prevention strategies.